Duffy Antigen Receptor For Chemokines Expression Research Articles

The role of Duffy antigen receptor for chemokines in keloids

This study aims to determine the relationship between Duffy antigen receptor for chemokines (DARC) and keloid pathogenesis. DARC expression was determined by immunohistochemistry, real-time PCR, and Western blot analysis. Cell proliferation was assessed by CCK-8 assay. Cell migration and invasion abilities were measured by the shift assay. Levels of CC chemokine ligand 2 (CCL2), CXC chemokine ligand 8 (CXCL8), and matrix metalloproteinase 2 (MMP2) were detected by real-time PCR and ELISA. Our results showed that DARC levels were elevated in human keloid fibroblasts. After knocking down DARC, cell proliferation was not altered, whereas the migration and invasion abilities of keloid fibroblasts were significantly elevated. Additionally, the mRNA expression levels of CCL2, CXCL8, and MMP2 were not influenced by DARC knockdown. However, the secretion of CCL2, but not CXCL8 or MMP2, was significantly increased after DARC knockdown. Our results suggest that DARC might inhibit the secretion of CCL2. Moreover, DARC knockdown increases the migration and invasion abilities of keloid fibroblasts.

Abstract P1-07-22: Investigating the role of Duffy antigen receptor for chemokines (DARC) isoforms in breast cancer metastasis

Abstract Duffy Antigen Receptor for chemokines (DARC) is known to regulate the biological activities of chemokines. It has been implicated in the progression of various diseases including different types of cancer, affecting survival, proliferation and metastasis. Genetic variants in the DARC genes are strongly associated with African ancestry and are linked to malaria resistance. There are at least two isoforms of DARC, which have not been appreciably investigated in these contexts, that may have differing chemokine activity functions. To fill this knowledge gap, we have been characterizing the impact of DARC gene variants on DARC isoform expression in specific tissue types in order to define relationships between the polymorphic DARC and its effect as an immune regulator in breast cancers. We hypothesize that DARC variants, which are known to impact expression of the gene on erythrocytes, also are associated with functional activity related to DARC’s control of metastatic potential in breast cancer. Specifically, we have determined that one DARC polymorphism which is well defined in ancestral groups (i.e. Duffy null - rs2814778; -541T>C) is associated with differential expression of DARC isoforms in epithelial cell types, including lymphoblast and breast tissues. We investigated the erythroid silencing Duffy Null allele of DARC using DNA sequencing and quantitated DARC isoform expression by real time PCR in breast cancer cell lines, ancestral cell lines and breast cancer tumor tissues. We will show that DARC isoforms have significantly variant expression among ancestral groups that is associated with the Duffy Null allele. We will also show that certain types of breast tumors have variable DARC expression that is also associated with this genotype. Additionally, we visualized the subcellular localization of DARC and known interacting chemokines breast cancer cells with IHC and IF confocal microscopy. Specifically, our results indicate breast tumors differentially express DARC in which may be affecting the chemokine pool and immune response within the tumor and its microenvironment. In our Hapmap ancestral populations, all DARC genotypes expressed the gene in lymphoblast, with significantly different levels of isoforms among African, African American and European individuals. Breast cancer cells also show different levels of specific DARC isoform mRNA levels and distinct levels and patterns of binding chemokines, related to tumor subtype origins (i.e. ER status). Based on these findings we conclude that DARC isoforms have differential expression levels in breast cancer cells within tumor and non-tumor regions that may drive both pro and anti tumorigenic events with varying chemokine binding or localizations during tumorigenesis of specific breast cancer subtypes. Different levels of isoforms in epithelial cells may contribute to its essential function as atypical chemokine receptor. We hypothesize that immune responses in breast cancer patients with Duffy Null genotypes will differ and may express more severe metastasis due to altered epithelial regulation of alternate DARC isoforms. Citation Format: Rupali Hire, Andrea Walens, Brianna Bennett, Kauthar Mumin, Hailey Campbell, Michael Lou, Michele Monteil, Elizabeth Howerth, Melissa Davis. Investigating the role of Duffy antigen receptor for chemokines (DARC) isoforms in breast cancer metastasis [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-07-22.

Studies of a murine monoclonal antibody directed against DARC: reappraisal of its specificity.

Duffy Antigen Receptor for Chemokines (DARC) plays multiple roles in human health as a blood group antigen, a receptor for chemokines and the only known receptor for Plasmodium vivax merozoites. It is the target of the murine anti-Fy6 monoclonal antibody 2C3 which binds to the first extracellular domain (ECD1), but exact nature of the recognized epitope was a subject of contradictory reports. Here, using a set of complex experiments which include expression of DARC with amino acid substitutions within the Fy6 epitope in E. coli and K562 cells, ELISA, surface plasmon resonance (SPR) and flow cytometry, we have resolved discrepancies between previously published reports and show that the basic epitope recognized by 2C3 antibody is 22FEDVW26, with 22F and 26W being the most important residues. In addition, we demonstrated that 30Y plays an auxiliary role in binding, particularly when the residue is sulfated. The STD-NMR studies performed using 2C3-derived Fab and synthetic peptide corroborated most of these results, and together with the molecular modelling suggested that 25V is not involved in direct interactions with the antibody, but determines folding of the epitope backbone.

Distribution of Duffy Antigen Receptor for Chemokines (DARC) and Risk of Prostate Cancer in Barbados, West Indies.

Blood typing across different racial groups has revealed that Caucasians predominantly test positive for the Duffy antigen/receptor for chemokines (DARC), while 70-95% of African-origin populations lack expression of DARC on their erythrocytes. Since men of African descent are known to have higher rates of prostate cancer (PC) and some animal studies have indicated anti-angiogenic effects associated with Duffy-positive mice, DARC-negativity may help to explain some of the racial differences in prostate tumorigenesis. The Prostate Cancer in a Black Population (PCBP) Study, a large case-control investigation including 1,007 incident PC cases and 1,005 controls, performed DARC testing on a subset of 1,295 participants (641 cases, 654 controls). The relationship between DARC expressivity and PC risk was evaluated using logistic regression models and findings are presented as odds ratios and 95% confidence intervals. More than three-quarters (76.5%) of African-Barbadian men lacked DARC expression, whereas almost three-fifths (59.3%) of White participants tested positive for the Duffy a and b alleles. DARC-negativity was not found to be associated with PC risk in the present investigation [OR 1.04, 95% CI (0.78, 1.37)], regardless of tumor grade. Findings from the PCBP study indicate that the majority of African-Barbadian men do not express DARC on their erythrocytes, yet absence of expression does not appear to be associated with PC development in this population.

Duffy antigen / receptor for chemokines correlates with inflammatory reaction in rats with venous hypertension: implication for the pathogenesis of primary chronic venous disease

Duffy antigen / receptor for chemokines (DARC) possesses high affinity for several chemokine subgroups of CC and CXC. Although DARC has been shown to play a role in many inflammatory diseases, its effect on chronic venous disease (CVD) remains unidentified. We explored whether the expression of DARC in skin tissue was activated under venous hypertension as well as the relationships between DARC and inflammation. The inflammation in a rat model of venous hypertension caused by a femoral arterial-venous fistula (AVF) was studied. At specified intervals the pressure in the femoral veins was recorded within 42 days. Hindlimb skin specimens were harvested at different time points. The expressions of DARC, interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) in skin tissue were examined. Mononuclear cells infiltrated in skin tissue were detected. Femoral venous pressures in AVF groups increased significantly at different time points (P < 0.01). DARC was expressed in skin tissue and its expression level increased significantly in AVF groups from the 7nd day on and was enhanced in a time-dependent manner within 42 days (P < 0.05). Meanwhile, both MCP-1 and IL-8 had higher levels, accompanied by increased mononuclear cells infiltrating into skin tissue (P < 0.05). A rat AVF model which can maintain venous hypertension for at least 42 days is competent for researching the pathogenesis of CVD. DARC, which plays a role in the inflammation of skin tissue under venous hypertension, may become a new molecular target for diagnosis and treatment of CVD at a very early stage.

The Development Of The Humanized Darc Transgenic Mouse

The Duffy Antigen Receptor for Chemokines (DARC) is an atypical chemokine receptor which binds CXC and CC chemokines with no measurable downstream signaling. Erythrocyte DARC serves as a chemokine sink and reservoir. DARC on endothelial cells transports chemokines thus promoting transendothelial leukocyte migration. Two common polymorphisms define the Duffy blood group, FYA and FYB, whilst the third polymorphism which is widely prevalent in West Africa, FYB(ES), abolishes erythrocyte expression of DARC. DARC is a portal of entry for plasmodium vivax suggesting that FYB(ES) was selected due to it conferring resistance to infection. The impact of the FYB(ES) polymorphism extends to a genetic association with the benign ethnic neutropenia. In order to study the mechanisms of this association we have developed a murine transgenic models of the Fy(a-b-) phenotype as encoded by FYB(ES), and Fy(a-b+) phenotype as encoded by FYB. Cloned genes were introduced by pronuclear micro-injection into fertilized oocytes of murine-DARC knockouts. The resulting embryos were transferred into pseudopregnant mice and offspring screened for transgene insertion by PCR genotyping. Gene expression in founder animals was measured by qPCR, demonstrating expression in lung, kidney, liver, spleen, skin and cerebellum. Protein expression was confirmed by immunofluorescence on frozen sections in spleen and skin in a distribution identical to human. Erythrocyte phenotyping by FACS using anti-Fy3 and anti-Fy6 monoclonal antibodies demonstrated the lack of DARC expression in mice with FYB(ES) and confirmed expression on FYB transgenic erythrocytes. We show that murine chemokines bind to human DARC and we plan to use this model to investigate the impact of FYB(ES) on in vivo chemokine functions and equilibria. These models will prove useful in the dissection of the mechanisms of ethnic neutropenia and improve the understanding in the role of DARC in chemokine homeostasis. Disclosures:Chaganti:Roche: Membership on an entity's Board of Directors or advisory committees, Receipt of travel grant Other.

Experimental study of Duffy antigen receptor for chemokines in tissue and the inflammatory reaction of limb with venous hypertension

To explore the role and mechanism of Duffy antigen receptor for chemokines (DARC) of tissue in promoting the inflammatory reaction of the limb with venous hypertension. moral arteriovenous fistula was surgically created to establish the rat model of venous hypertension. A total of 36 SD rats were randomly divided into pcDNA3.1-DARC (Group A), empty plasmid of pcDNA3.1 (Group B) and control (Group C) groups. The animals were sacrificed at Days 14 and 42 post-operation respectively. The expressions of DARC at the RNA and protein level were detected by real-time polymerase chain reaction (PCR) and Western blot. And the serum level of interleukin (IL)-8 was detected by enzyme linked immunosorbent assay (ELISA) and the degrees of apoptosis and leukocytic infiltration of local tissue were detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) and hematoxylin and eosin (HE) staining. With the elapsing time of venous hypertension, the DARC expression in tissue, the extent of apoptosis and leukocytic infiltration in tissue showed an increasing trend in Groups A and B. Group A was obviously higher than Group B during the corresponding period. And the differences were statistically significant (P < 0.05). The serum levels of IL-8 of Groups A and B showed a decreasing trend. And Group A was obviously lower than Group B. Both groups were higher than the control group. The differences were statistically significant (P < 0.05). The level of DARC in tissue and the degree of inflammatory reaction of venous hypertension have a positive correlation. And DARC may promote the development of venous hypertension inflammation through augmenting the adhesion and migration of leukocytes.

Duffy antigen receptor for chemokines in laryngeal squamous cell carcinoma as a negative regulator

Conclusion: Duffy antigen receptor for chemokines (DARC), as a negative regulator, could attenuate the potential growth and metastasis of laryngeal squamous cell carcinoma (LSCC); hence it could be considered as a basis for a new prognostic aid and molecular therapeutic target in treating LSCC. Objectives: To analyze the role of DARC in LSCC progression using LSCC clinical samples. Methods: A total of 65 LSCC tissues were examined immunohistochemically for DARC protein expression. The microvessel density (MVD) was determined by the endothelial cells immunostained using anti-CD34 antibody. Results: It was found that DARC was expressed in the LSCC samples, with a high expression in 32 of our cases (49.2%), and in lymph node-negative and -positive groups of 24 (72.7%) and 8 cases (25%), respectively (p < 0.01). MVD was (24.75 ± 5.45)/High power field (HP) and (32.58 ± 5.23)/HP in the cases with high and low expression of DARC, respectively (p < 0.01). More importantly, the DARC expression presented a strong negative correlation with nodal metastasis, with pTNM stage, and with recurrence, and the cases with a high DARC expression indicated a higher survival rate. In addition, the multivariate analysis showed that the DARC expression was an independent prognostic factor (p = 0.032; hazard ratio, 21.476; 95% confidence interval, 1.297–355.68).

Acquired Antibody Responses against Plasmodium vivax Infection Vary with Host Genotype for Duffy Antigen Receptor for Chemokines (DARC)

BackgroundPolymorphism of the Duffy Antigen Receptor for Chemokines (DARC) is associated with susceptibility to and the severity of Plasmodium vivax malaria in humans. P. vivax uses DARC to invade erythrocytes. Individuals lacking DARC are ‘resistant’ to P. vivax erythrocytic infection. However, susceptibility to P. vivax in DARC+ individuals is reported to vary between specific DARC genotypes. We hypothesized that the natural acquisition of antibodies to P. vivax blood stages may vary with the host genotype and the level of DARC expression. Furthermore, high parasitemia has been reported to effect the acquisition of immunity against pre-erythrocytic parasites. We investigated the correlation between host DARC genotypes and the frequency and magnitude of antibodies against P. vivax erythrocytic stage antigens.Methodology/FindingsWe assessed the frequencies and magnitudes of antibody responses against P. vivax and P. falciparum sporozoite and erythrocytic antigens in Colombian donors from malaria-endemic regions. The frequency and level of naturally-acquired antibodies against the P. vivax erythrocytic antigens merozoite surface protein 1 (PvMSP1) and Duffy binding protein (PvDBP) varied with the host DARC genotypes. Donors with one negative allele (FY*B/FY*Bnull and FY*A/FY*Bnull) were more likely to have anti-PvMSP1 and anti-PvDBP antibodies than those with two positive alleles (FY*B/FY*B and FY*A/FY*B). The lower IgG3 and IgG1 components of the total IgG response may account for the decreased responses to P. vivax erythrocytic antigens with FY*A/FY*B and FY*B/FY*B genotypes. No such association was detected with P. falciparum erythrocytic antigens, which does not use DARC for erythrocyte invasion.Conclusion/SignificanceIndividuals with higher DARC expression, which is associated with higher susceptibility to P. vivax infection, exhibited low frequencies and magnitudes of P. vivax blood-stage specific antibody responses. This may indicate that one of the primary mechanisms by which P. vivax evades host immunity is through DARC indirectly down-regulating humoral responses against erythrocytic invasion and development.

The Duffy Antigen/Receptor for Chemokines (DARC) and Prostate-Cancer Risk among Jamaican Men

As an evolutionary response to prevent malaria infection, most Africans do not express the Duffy Antigen/Receptor for Chemokines (DARC) on their red blood cells. Results from experimental studies suggest that DARC expression inhibits prostate-tumor growth. We tested the hypothesis that men of African descent who lack DARC expression are at increased risk of prostate cancer. A case-control study involving 81 age-matched pairs was conducted in Jamaica. Participants were interviewed to collect data, and they donated blood for determination of DARC expression. Logistic regression was used to estimate associations with prostate cancer and aggressive disease. Little or no association was observed between erythrocyte DARC expression and prostate cancer or between DARC expression and aggressive disease. These associations changed little when adjusting for other potential confounders. Our results do not support an effect of erythrocyte DARC expression on the risk or progression of prostate cancer in men of African descent.

Association between Duffy antigen receptor for chemokines expression and levels of inflammation markers in sickle cell anemia patients

Since inflammation plays a prominent role in the pathogenesis of sickle cell anemia (SCA) and Duffy antigen receptor for chemokines (DARC) modulates the function of inflammatory processes, we analyzed the relationship between the erythrocyte DARC phenotype and clinical expression of SCA. DARC locus was genotyped in 212 SS adult patients followed by the sickle cell center of Guadeloupe (French West Indies). After patients' stratification according to RBC DARC expression, the prevalence of renal disease, leg ulcers, priapism and osteonecrosis was compared between patient groups as well as hematological variables and plasma levels of chemokines. Duffy-positive patients exhibited higher counts of white blood cells (9.95+/-2.36 vs 8.88+/-2.32 10(9)/L, p=0.0066), polynuclear neutrophils (5.1+/-1.73 vs 4.51+/-1.71 10(9)/L, p=0.0227), higher plasma levels of IL-8 (4.46+/-1.22 vs 1.47+/-0.5 pg/mL, p=0.0202) and RANTES (27.8+/-4.3 vs 18.1+/-2.3 ng/mL, p=0.04) than Duffy-negative patients. No association was detected between RBC expression of DARC and the studied complications.

The role of chemokine receptors in acute lung allograft rejection

Recruitment of inflammatory cells to vascularised allografts is a hallmark of rejection, and paves the way for chronic allograft injury. Chemokines play pivotal roles in the directed movement of leukocytes. Herein, we define the distribution of chemokine receptors for the most common cell types during human lung allograft rejection as a prerequisite for therapeutic interventions. Immunohistochemistry was performed on lung allograft biopsies from 54 patients for the chemokine receptors CCR5, CXCR3 and CXCR1 and the Duffy antigen/receptor for chemokines (DARC). Perivascular infiltrates in acute lung rejection are composed of subsets of mononuclear cells expressing the chemokine receptors CXCR1, CXCR3 and CCR5. DARC-positive small vessels and capillary vessels were associated with sites of inflammation and their number was increased during episodes of acute lung rejection. DARC expression correlated with an increase in interstitial CCR5-positive T-cells and CXCR1-positive leukocytes. Leucokytic infiltrates in bronchial/bronchiolar rejection express CXCR1 and CXCR3. This is the first study that demonstrates an induction of the chemokine binding protein DARC at sites of acute human lung allograft rejection. Co-localisation with the chemokine receptors CXCR1 and CCR5 may indicate a role for DARC expression during leukocyte adhesion and interstitial infiltration.

Shedding light on DARC: the role of the Duffy antigen/receptor for chemokines in inflammation, infection and malignancy

Duffy antigen (Fy) was first identified in 1950 as a cause of transfusion reactions [1], and 25 years later recognized as receptor allowing the entry of Plasmodium vivax into erythrocytes [2]. It was not until the mid 1990s that a physiological role for this antigen was proposed, when Duffy was found to be a chemokine receptor [3] and was subsequently renamed Duffy antigen receptor for chemokines (DARC). Since then roles for DARC in inflammation, infection and malignancy have begun to emerge. Most individuals from Sub-Saharan Africa, or of African descent, have a mutation that prevent DARC expression on erythrocytes, but not other sites of DARC expression, such as endothelial cells. Interest in the clinical and scientific implications of DARC has risen as it has become a potential explanation for some of the differences in prognosis and presentation seen in those with African genealogy in diseases of inflammation and malignancy. This paper reviews the evidence for DARC influencing inflammation, and its role in diseases of inflammation, infection and malignancy. DARC as a regulator of inflammation

Duffy antigen receptor for chemokines and CXCL5 are essential for the recruitment of neutrophils in a multicellular model of rheumatoid arthritis synovium.

The role of chemokines and their transporters in rheumatoid arthritis (RA) is poorly described. Evidence suggests that CXCL5 plays an important role, because it is abundant in RA tissue, and its neutralization moderates joint damage in animal models of arthritis. Expression of the chemokine transporter Duffy antigen receptor for chemokines (DARC) is also up-regulated in early RA. The aim of this study was to investigate the role of CXCL5 and DARC in regulating neutrophil recruitment, using an in vitro model of RA synovium. To model RA synovium, RA synovial fibroblasts (RASFs) were cocultured with endothelial cells (ECs) for 24 hours. Gene expression in cocultured cells was investigated using TaqMan gene arrays. The roles of CXCL5 and DARC were determined by incorporating cocultures into a flow-based adhesion assay, in which their function was demonstrated by blocking neutrophil recruitment with neutralizing reagents. EC-RASF coculture induced chemokine expression in both cell types. Although the expression of CXC chemokines was modestly up-regulated in ECs, the expression of CXCL1, CXCL5, and CXCL8 was greatly increased in RASFs. RASFs also promoted the recruitment of flowing neutrophils to ECs. Anti-CXCL5 antibody abolished neutrophil recruitment by neutralizing CXCL5 expressed on ECs or when used to immunodeplete coculture-conditioned medium. DARC was also induced on ECs by coculture, and anti-Fy6 antibody or small interfering RNA targeting of DARC expression effectively abolished neutrophil recruitment. This study is the first to demonstrate, in a model of human disease, that the function of DARC is essential for editing the chemokine signals presented by ECs and for promoting unwanted leukocyte recruitment.

Opposing roles of murine duffy antigen receptor for chemokine and murine CXC chemokine receptor-2 receptors in murine melanoma tumor growth.

The Duffy antigen receptor for chemokines (DARC) has been classified as a "silent" receptor, as it can bind CXC and CC chemokines to undergo ligand-induced receptor internalization, but is not coupled to trimeric G proteins required for the classic G protein-coupled receptor-mediated signaling. CXC chemokine receptor-2 (CXCR2) has been shown to play a major role in tumor angiogenesis. To test the hypothesis that these two chemokine receptors might play opposing roles in the growth of melanoma tumors, we developed a transgenic mouse model, where the preproendothelin promoter/enhancer (PPEP) is used to drive expression of either murine DARC (mDARC) or murine CXCR2 (mCXCR2) in endothelial cells. We show herein that the growth of melanoma tumor xenografts, established from s.c. injection of immortalized murine melanocytes overexpressing macrophage inflammatory protein-2, was inhibited or enhanced in the PPEP-mDARC and PPEP-mCXCR2 transgenic mice, respectively, compared with control mice. The early tumors formed in mDARC transgenic mice exhibited a significantly higher number of infiltrating leukocytes compared with either the control or mCXCR2 transgenic mice, suggesting a potential role for DARC expressed on endothelial cells in leukocyte migration. In addition, the tumor-associated angiogenesis in mDARC transgenic mice was reduced when compared with the control. Conversely, tumor angiogenesis was significantly increased in mCXCR2 transgenic mice. Results indicate that endothelial cell overexpression of mDARC increased leukocyte trafficking to the tumor, reduced the growth of blood vessels into the tumor, and reduced the growth rate of the tumor, whereas endothelial cell overexpression of mCXCR2 had the reverse effect on tumor angiogenesis and growth.

Impact of Erythrocyte Duffy Antigen Genetic Polymorphism on the Distribution of GroB-T, a Novel Human CXC Chemokine

Grobeta-T, a human CXC chemokine, has been studied for its potential to mobilize stem cells. Chemokines bind specifically to receptors on target immune cells but also to a homologous erythrocyte blood group antigen, the Duffy Antigen/Receptor for Chemokines (DARC)that is subject to genetic polymorphism in humans.A mutation in the DARC gene is common among African Americans and results in lack of expression of the erythrocyte antigen. We used a combination of in vitro studies of Grobeta-DARC interaction and pharmacokinetic simulation to anticipate the potential impact of this polymorphism on the pharmacokinetics of Grobeta-T. [125I]Grobeta-T was incubated in Caucasian blood to characterize the concentration dependence of the blood to plasma concentration ratio (B/P). Affinity and capacity of binding was estimated by Scatchard analysis; specificity was investigated by competitive displacement with a CC chemokine. The B/P value (7 nM) was then determined in blood from 8 African American subjects. Duffy antigen expression was determined by antibody agglutination. A pharmacokinetic model was developed which accounted for blood-cell binding. Simulations were performed to explore effects of dose regimen and DARC expression on the Grobeta-T plasma concentration-time profile. Grobeta-T affinity and capacity for DARC (Caucasian blood)were 23.0 +/- 1.2 and 37.7 +/- 0.6 nM, respectively;excess CC chemokine fully displaced [125I]Grobeta-T. Chemokine binding was highly correlated with the presence or absence of the Duffy antigen (p<0.01) in African American blood; the proportion of subjects for which binding was observed (3/8), was consistent with the reported frequency of DARC expression in this population. Counter to intuition,in the terminal disposition phase at low doses,concentrations of free Grobeta-T in the presence of DARC may be substantially higher than in the absence of DARC. Dissociation from the erythrocyte antigen may lead to greater persistence, at low doses, of free Grobetabeta-T in the blood of individuals expressing the chemokine sink.

Duffy antigen receptor for chemokines (DARC) on non‐hematopoietic tissues curbs atherosclerosis in apolipoprotein E‐deficient mice

The Duffy antigen receptor for chemokines (DARC) is a non-signaling, promiscuous chemokine receptor expressed on red blood cells (RBCs) and high endothelial venules (HEVs). The function of RBC DARC and HEV DARC in inflammatory disease is not well understood. In order to examine the role of DARC in atherosclerosis, we backcrossed apolipoprotein E knockout (apoE−/−) mice with DARC knockout (DARC−/−) mice to create double knockout (apoE−/−/DARC−/−) mice. apoE−/−/DARC−/− and apoE−/− mice were fed a Western diet for 12 weeks. Aortas were then stained with Oil Red O and percent aortic area with atherosclerotic lesion was quantified. apoE−/−/DARC−/− and apoE−/− mice developed lesions on 6.1 ± 2.7% and 4.2 ± 1.4% (average ± SEM, p<0.05) of their aortas, respectively. To differentiate between RBC DARC and HEV DARC, we performed the same analysis on apoE−/− mice and apoE−/−/DARC−/− mice reconstituted with either apoE−/− or apoE−/−/DARC−/− bone marrow. DARC expression on non-hematopoietic cells protected mice from atherosclerosis to the same extent as complete DARC expression (4.6 ± 1.0% and 4.9 ± 1.1%, respectively, p>0.05). By contrast, apoE−/−/DARC−/− receiving either apoE−/− (8.3 ± 1.6%) or apoE−/−/DARC−/− (7.8 ± 1.7%) bone marrow exhibited a 1.5-fold increase in lesion areas compared to mice expressing DARC on non-hematopoietic cells (p<0.05 for groups expressing DARC on non-hematopoietic tissue versus those deficient in DARC on non-hematopoietic tissue). We conclude that HEV DARC, but not RBC DARC, is involved in curbing atherosclerosis. This work was supported by NIH HL 58108 and the American Heart Association.

The Duffy antigen/receptor for chemokines (DARC) regulates prostate tumor growth.

The Duffy antigen/receptor for chemokines (DARC) is a promiscuous chemokine receptor that binds to members of the CXC chemokine family possessing angiogenic properties. The DARC is expressed on erythrocytes and endothelial cells and is required for Plasmodium vivax infection of erythrocytes. Approximately 70% of African-Americans lack erythrocyte expression of the DARC as a genetic mechanism of protection against malaria infection. African-American men have a 60% greater incidence of prostate cancer and a 2-fold higher mortality rate than Caucasian men. Using a transgenic model of prostate cancer with DARC-deficient mice, we tested the hypothesis that lack of DARC expression on erythrocytes contributes to enhanced prostate tumor growth. In vitro, erythrocytes from wild-type mice but not DARC-deficient mice cleared angiogenic chemokines produced by prostate cancer cells and reduced endothelial cell chemotaxis. In vivo, tumors from DARC-deficient mice had higher intra-tumor concentrations of angiogenic chemokines, increased tumor vessel density, and greatly augmented prostate tumor growth. The data suggest that the DARC functions to clear angiogenic CXC chemokines from the prostate tumor microcirculation and that the lack of erythroid DARC, as occurs in the majority of African-Americans, may be a contributing factor to the increased mortality to prostate cancer in this population.

Duffy Antigen/Receptor for Chemokines (DARC): Is There a Role in Prostate Cancer?.

Introduction: The Duffy blood group antigens function as chemokine receptors and as receptors for several species of malarial parasites. Approximately 70% of African Americans are Duffy negative as a result of a single nucleotide polymorphism in the promoter region of the Duffy antigen/receptor for chemokines (DARC) gene in a consensus-binding site for GATA 1. This mutation abolishes the promoter activity in erythrocytes, impairing the expression of DARC on red blood cells. Other than resistance to certain species of malaria, the functional consequence of Duffy negativity is unclear; however, it has recently been proposed that the Duffy antigen acts as a biological ‘sink' to clear pro-inflammatory chemokines from tissue microcirculation. Further it has been suggested that since the incidence of prostate cancer in African Americans is 60% higher than in Caucasians, absence of the Duffy antigen might predispose African Americans to prostate cancer by impairing downregulation of proinflammatory cytokines. We tested the hypothesis that lack of expression of DARC on erythrocytes predisposes African American men to develop prostate cancer.Methods: We conducted a case control study of 89 African American men with confirmed prostate cancer and 51 age matched healthy African American men. The samples were genotyped for the promoter polymorphism using an allele specific real time PCR assay (Taqman assay) developed for this study.Results: The frequency of the Duffy negative allele was 75.8% among the patients and 81.4% among the controls (odds ratio = 0.72, p value = 0.28). The distribution of heterozygous and homozygous Duffy negative patients did not differ between the cases and controls.Conclusion: It has been suggested that individuals lacking erythrocyte expression of DARC have higher prostate levels of angiogenic chemokines that might promote more rapid development of prostate cancer. The data presented here found no difference in the frequency of the Duffy negative allele between African American men with prostate cancer and healthy age matched African American controls. Since several studies suggest that polymorphisms associated with differential production of IL-8, IL-10 and VEGF are risk factors for prostate cancer, it remains possible that a polymorphism associated with differential cytokine production in combination with Duffy negativity increases the risk for prostate cancer.

Overexpression of the duffy antigen receptor for chemokines (DARC) by NSCLC tumor cells results in increased tumor necrosis.

BackgroundThe Duffy antigen receptor for chemokines (DARC) is known to be a promiscuous chemokine receptor that binds a variety of CXC and CC chemokines in the absence of any detectable signal transduction events. Within the CXC group of chemokines, DARC binds the angiogenic CXC chemokines including IL-8 (CXCL8), GROα (CXCL1) and ENA-78 (CXCL5), all of which have previously been shown to be important in non-small cell lung carcinoma (NSCLC) tumor growth. We hypothesized that overexpression of DARC by a NSCLC tumor cell line would result in the binding of the angiogenic ELR+ CXC chemokines by the tumor cells themselves, and thus interfere with the stimulation of endothelial cells and induction of angiogenesis by the tumor cell-derived angiogenic chemokines.ResultsNSCLC tumor cells that constitutively expressed DARC were generated and their growth characteristics were compared to control transfected cells in vitro and in vivo in SCID animals. We found that tumors derived from DARC-expressing cells were significantly larger in size than tumors derived from control-transfected cells. However, upon histological examination we found that DARC-expressing tumors had significantly more necrosis and decreased tumor cellularity, as compared to control tumors. Expression of DARC by NSCLC cells was also associated with a decrease in tumor-associated vasculature and a reduction in metastatic potential.ConclusionsThe expression of DARC in the context of NSCLC tumors may act as a chemokine decoy receptor and interferes with normal tumor growth and chemokine-induced tumor neovascularization.